| The potato products processing and starch industries produce waste effluent of potato fruit juice containing substantial amounts of protein.Patatin is one the major component of this industrial effluent with high nutritional value and profound functional properties.The study has shown the potential of potato patatin.In this study,the effect of high hydrostatic pressure?HHP?on potato patatin was investigated and its physicochemical characteristics and functional properties were explored.Moreover,the effect of HHPtreatment on structural modification,and its antioxidant potential and iron chelation activities were assessed.Finally,the anticancer activities of HHP treated patatin onhuman colon adenocarcinoma?HT-29?cells were investigated invitro.Patatin?40 kDa?was extracted and precipitated using ammonium sulfate?AS?from potato fruit juice and further purified by ion exchange chromatography followed by gel chromatography,of which the structural characteristics were investigated.Purified patatin wastreated by HHP?250,350,450,and550 MPa?.SDS-PAGE results confirmed the MW of patatin was 40 kDa in bothnative and HHP-treated patatin.HHP-treated patatin?250–550 MPa?showed a clearly increasing pattern of new protein bands at MW>130 kDa,suggesting that aggregation occurred due to HHP treatment.HHP treatment also effected monosaccharide content of patatin,including arabinose,rhamnose,galactose,glucose and xylose.There was a significant increase in galactose content,which was 18.04%and 17.03%at 250MPa and 550 MPa,respectively;whereas a subsequent decrease in glucose contentwas observed afterHHP treatment,which was 10.93%,10.70%,12.30%and 11.24%from 250 to 550 MPa,respectively.FTIR and CD spectra briefed prominent changes in HHP-treatedpatatin secondary structure as compared with native patatin?NP?.FTIR results showed significant increases in transmittance intensity along with increases in pressure from 250 to 550 MPa.Furthermore,HHP treatment influenced the intermolecular and intramolecular attractions in unfolded protein structures by increasing?-sheet conformations and reductions in?-helices.Thermal changes observed by differential scanning calorimetry?DSC?also showed a similar trend following HHP treatment;however,the enthalpy of patatin was negatively affected by pressurization.Surface hydrophobicity and free sulfhydryl content significantly?p<0.05?increased with pressurization up to 450 MPa,and marked increase in free-SH groups might be related to the unfolding of the protein following exposure of embedded SH groups.Moreover,the antioxidant potential and iron chelation activities of HHP-treated patatin were enhanced significantly.HHP-treated?550 MPa?patatinshowedthe highest oxygen radical absorbance capacity?ORAC?,which was 110,700?M TE 100 g-1 at 4 mg mL-1.The effect of HHP on anticancer activity of patatin on human colorectal cancer?HT-29?cell was investigated.The NP and HHP-treated patatin inhibited cell proliferation withdose dependant manner.Compared with NP,the proliferation inhibition of HHP-treated patatin on HT-29 cellsenhanced significantly,which was the highest when treated at 450 MPa.AO/EtBr staining revealed chromatin condensation,fragmentation and apoptotic bodies in HT-29 cells.The percentage of late apoptotic and necrotic cells were in HHP-treated?450 MPa?patatinthen followed by treatment at 550 MPa.The autophagic assay by neutral red staining indicated increased autophagic cells with increasing pressurization level.The 3 mg mL-1 concentration of both HHP-treated patatin at 450 and 550 MPa,showed maximum number of neutral red stained bodies?NRSBs?in HT-29 cells indicating increased autophagy compared to the same concentration of NP and patatin pressurized at 250 and 350 MPa.DNA fragmentation appeared in NP treated cellswith less intensity as compared to HHP-treated patatin.Cells incubatedwith HHP-treated patatin at 350,450 and 550 MPa showed band boarding and smearing of DNA represented the induction of apoptosis under the effect of pressurized patatin.HHP treatment offers an effective and green process for inducing structural modifications and improving patatin functionality. |
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